A growing body of studies developing and investigating three-dimensional scaffolds have proven that biophysical parameters are an essential factor necessary to mimic the cellular environment, allowing for the regeneration of tissues and organs [1, 2]. While mechanically supporting cellular environments, three-dimensional scaffolds are often designed with porous and interconnected networks that guide cellular behaviors, resulting in successful tissue and organ regeneration [3]. Porous structures within a biomaterial are capable of nurturing, growing, and differentiating cells, contributing to improved tissue regeneration [4]. In the field of tissue engineering, there have been many attempts to make three-dimensional scaffolds that allow cell growth and tissue regeneration. For example, solvent casting, particle leaching, gas foaming, and melt molding have all been proposed. Additionally, rapid prototyping systems have also been considered in order to control the size, pore geometry, and interconnectivity of scaffolds. Common rapid prototyping systems utilize selective laser sintering (SLS), fused deposition modeling (FDM), stereolithography (SLA), 3D printing as well as 3D plotting to generate scaffolds [5-10]. However, one of the most studied and popular methods for fabricating scaffolds is electrospinning. Electrospinning has been widely applied to the fabrication of biomimetic scaffold that remodels the native extracellular matrix (ECM) [11]. Conventional electrospinning is an easy and commonly utilized method for the fabrication of nanofibrous polymer scaffolds that are capable of supporting cell growth.
Traditional electrospinning employs a conductive collector that is set some distance away from a syringe containing a polymer solution. Both the syringe and the collector are hooked up to a high voltage source, with the positive charge connected to the syringe and the ground connected to the collector, generating a voltage differential. When a high voltage is applied to the collector and the syringe, the polymer overcomes the surface tension of the syringe and is deposited on the collector in a nanofiber configuration. The resulting electrospun scaffolds are composed of nanofibrous layers arranged in a tightly packed conformation.
However, traditional electrospinning methods are limited in the sense that they are not capable of fabricating porous scaffolds to allow the infiltration of cells without additional modification. Many attempts have been made to make a porous scaffold with consistency, but the majority of these methods are labor intensive and do not result in consistent porous structures. In fact, most methods require extra handling steps and produce patterning and porosity that is unpredictable and random. For example, one such method involves the incorporation of a water soluble material such as salt into the polymer solution. During the electrospinning process, the salt is deposited into the polymer fibers and incorporated into the scaffold. Following formation of the nanofibrous sheet, it is submerged into water in order to leach the salt out, leaving behind porous defects [12]. However, these porous defects are not interconnected throughout the scaffold and are simply pouches that allow cell populations to form enclaves within the scaffold. Furthermore, these porous defects do not allow cells to grow throughout the scaffold.
One example where electrospun scaffolds may be beneficial is in guided bone regeneration (GBR). GBR is a clinically proven technique used to restore maxillofacial defects using barrier membranes to cover the defects and induce bone regeneration [13]. In this regard, the membrane is intended to prevent soft-tissue ingrowth into the bone defect while promoting bone tissue regeneration [14, 15]. The osteoinductive space created by a membrane sheet enables bone formation around the periodontal defect with successful prevention of gingival connective tissue invasion, which can hinder osteoinduction. Dahlin et al. used a porous polytetrafluoroethylene (PTFE) membrane to facilitate the migration of cells responsible for osteogenesis to the defect site within the mandibular angles of Sprague-Dawley rats [16]. Although PTFE's biological inertness is a beneficial property for vascular grafts, the use of such non-resorbable materials often leads to postoperative healing complications [17]. For this reason, a second surgical manipulation to remove the non-resorbable membrane after 4 to 6 weeks following the initial operation is unavoidable.
Unlike traditional PTFE membranes, resorbable membranes do not require a secondary surgical manipulation, leading to successful GBR with reduced patient discomfort. In general, two types of materials have been studied for use as resorbable membranes: synthetic resorbable membranes and naturally biodegradable membranes. Poly(glycolic acid) (PGA), poly(lactic acid) (PLA), poly(ε-caprolactone) (PCL), and their copolymers are the most commonly used materials for synthetic resorbable membranes, whereas bovine and porcine collagen membranes are commercially available, naturally biodedgradable materials used for GBR [18, 19, 20]. All resorbable membranes have previously been modified and reinforced in order to meet essential features necessary for successful GBR. These features include biocompatibility, selective cell ingrowth for bone regeneration, space maintenance, mechanical stability, adequate degradability, good tissue integration, and ease of use [21]. Of these characteristics, selective cell ingrowth following migration is a critical parameter necessary to facilitate the migration of osteogenic cells while retarding the ingrowth of gingival connective tissue. This feat is usually achieved via the utilization of a multi-layered membrane [22]. For instance, Epi-Guide®, which is a porous membrane made from three layers of a poly-D, DL, L-lactic acid polymer, retains fibroblasts and epithelial cells within the membrane while the layered barrier membrane allows bone regeneration by maintaining space around the defect up to 20 weeks. Another example is Bio-Gide®, which is a bi-layered membrane made of a non-cross-linked collagen membrane. The compact layer of this membrane provides a protective barrier for adjacent connective tissue while the porous layer supports bone and periodontal regeneration. Hence, for successful GBR, the generation of a porous structure without sophisticated methodologies would be preferred. Ideally, a technique that enables the production of a selective, patterned porous structure within the multi-layered membrane would have great potential for use in resorbable membranes for GBR. Thus, this invention involves a novel electrospinning technique in which a modified collector apparatus is used to provide a much less demanding process for the fabrication of a uniformly patterned and porous nanofibrous structure within a resorbable GBR membrane.